Defoaming process



2,759,871 Patented Aug. 21, 1956,

DEFOAMING PROCESS Jacob S. Cash, Syracuse, N. Y., assignor toBristol'LaboYratltlries Inc., Syracuse, N. Y., a corporation of New NoDrawing. Application August 6, 1952, Serial No. 303,008

This invention relates in general'to a process and means for producingchemicals by the deep culture, aerated fermentation method, and moreparticularly to av process and means for abating and reducing foam inthe production of chemicals, including antibiotics, by the deep culturefermentation method.

Numerous chemicals, including antibiotics in particular, are producedcommercially by growing an organism, e. g. bacteria, fungi, in largetanks containing nutrient materials while passing in a large amount ofair. During this time excessive foaming frequently occurs. This reducesmarkedly the actual. production in a given tank because the fermentationtank cannot be utilized to full capacity. The non-toxic defoaming agentspresently available are often not sufiiciently effective to permit useof the full capacity of the tank even when large quantities of defoameragent are used, c. g. as much as one or two percent of the weight of themedium in the tank. In addition, the use of large quantities of defoameragent renders more difiicult the isolation of the final product.

it is an object of the present invention to provide an improvement inthe process of producing chemicals and antibiotics, e. g. penicillin,streptomycin, by the deepculture, aerated fermentation method whichcomprises the step of abating and reducing foam therein by introducingdefoamer agent substantially below the surface of the medium as anaerosol in a stream of sterile, non-toxic gas.

It is a further object of the present invention to provide animprovement in the process of producing chemicals by the deep-culture,aeratedferrnentation method which comprises the step of abating andreducing foam therein by introducing defoamer agent substantially belowthe surface of the medium in a stream of non-toxic gas as a dispersionof particles having diameters of less than 1000 microns and preferablyin which 90% of the particles have diameters of less than 200 microns.

It is a further object of the present invention to pro.- vide animprovement in the process of producing antibiotics, e. g. penicillin,streptomycin, by the deep-culture, aerated fermentation method whichcomprises the step of abating and reducing foam therein by introducingdefoamer agent substantially below the surface of the me dium in astream of sterile, non-toxic gas as a dispersion of particles havingdiameters of less than 1000 microns and preferably in which 90% of theparticles have diameters of less than 200 microns.

It is a further object of the present invention to provide animprovement in deep-culture, aerated fermentation processes which makespossible a reduction in the. con-- sumption of material, i. e., defoameragent, and also reduces loss of the product via foaming.

There is now discovered according to the present invention, theimprovement in the process of producing, chemicals by the deep-culture,aerated fermentation method which comprises the step of abating'v andreducing foam therein by introducing defoamer agent substantially be- 2low the surface'of 'the medium as an aerosol in a stream of non-toxicgas.

A- more comprehensive understanding of this invention is obtained byreference to the following examples.

Example I Three duplicate fermentations identified as Runs 1, 2,- and 3were conducted simultaneously in tanks having a nominal capacity of 600gallons. media were those conventional in the prior art. To 325 gallonsof sterile lactose-calcium carbonate-corn steepphenylacetic acid mediawith added trace minerals was added as inoculum 65 gallons ofa-Penicillium fermentation. Fermentation was conducted at about F. withthe usual aeration and agitation for about 80 hours. At' the end of thefirst twenty-two hours, 60-additional gallons of original, sterilemedium were added. Lard oil containing one percent octadecanol was usedas the defoaming agent and the only variation between these three runswas in the manner of its addition.

In Runs 1 and 2, the'defoamer was dropped onto the surface of the brothin approximately cc. portions by an automatic device (spark plug) as thefoam reached a certain height in thetank. This amount of defoamer (100cc.) had previously been determined to be the minimum satisfactoryquantity. Approximately 75% of the consumption of the defoamer occurredin the first twenty hours. Foaming from about the sixth to tenth hourwas so great, despite the defoamer agent, that at times it was necessaryto reduce aeration and stop agitation. Both of these actions are highlydetrimental to the yield of penicillin.

In Run 3, the defoamer was aerosolized into the sparger air, by means ofcompressed air and a pneumatic atomizing nozzle of the type used forspraying paint, by manual operation as called for by an automatic device(spark plug) indicating that the foam had reached a certain height inthe tank. Each addition used about ten to fifteen cc. of defoamer agent.The apparatus in use passed about 340 cc./minute of lard oil at 50pounds per square inch gage pressure, using about 15 to 20 cubic feetper minute of air. Foaming during the entire fermentation was controlledso well that there was no need at any time to reduce aeration or to stopagitation.

The consumption of defoamer agent in pounds in Run 3 (2.5 pounds) was4.8 percent of the average consumption in Runs 1 and 2. The peak assayof the broth in Run 3 was 78% of the average peak assay in Runs 1 and 2.

Use of the defoamer agent in aerosol form gave slightly better controlof foam and markedly decreased the consumption of defoamer agent. Asseen by inspection of Example II, below, the lower titer of Run 3 may beattributed either to the usual variation of 20 percent encountered inbiological and biochemical work or to the fact that a minimum weight oflard oil of about one-tenth of one percent. of the weight of medium (i.e., four pounds in this case) is generally necessary to providesufficient esters of long-chain fatty acids as growth stimulantsaccording to Kofiier (Ind. and Eng. Chem, 42, 1819-1823 (1950) ExampleII Three duplicate fermentations identified as Runs 4, 5, and 6 wereconducted simultaneously in the manner of Example I. The only variationbetween the runs Was in the manner of addition of the defoamer agent; inRuns 4 and 5, as in Runs 1 and 2, the defoamer agent was dropped ontothe surface of the broth and in Run 6, as in Run 3, the defoamer agentwas added to the sparger air in aerosol form.

The consumption of defoamer agent in pounds in Run 6 (3.9 pounds) was13.7 percent of the. average consump- The techniques and" tion in Runs 4and 5. The peak assay of the broth in Run 6 was 98.5 percent of theaverage peak assay in Runs 4 and 5.

In Runs 4 and 5, but not in Run 6,.it was necessary to reduce aerationand stop agitation at times during the sixth to tenth hours because thefoaming was out of control.

Use of the defoamer agent in spray form gave slightly better control offoam and markedly decreased the consumption of defoamer agent withoutdecreasing the yield of penicillin.

As defoamer agents, octadecanol in lard oil or corn oil is preferred butuse may be made of any of the agents which have been found to beeffective and non-toxic in the surface drip method. These include, forexample, corn oil (contains approximately 46.3% oleic, 42% linoleic,7.5% palmitic, 3.5% stearic and traces of arachidonic and lignocericacid), Defoamer (Swift and Co., contains lard oil, mineral oil, and asurface-active fatty acid derivative), Defoamer 51 (Swift and Co.,contains a fat more saturated than lard oil, mineral oil and asurface-active fatty acid derivative), lard oil (Swift and Co., containsapproximately 65% oleic, linoleic, 2% lino-lenic and 23%stearic-palmitic acid), Monosulph (Nopco Chemical Co., sulfonated oilcontaining oleic and hydroxyoleic acids), Nopalcol 1-0 (Nopco ChemicalCo., mono-oleic acid ester of glycol), Nopco glyceryl oleate (mixture ofmonoand di-oleic acid esters of glycerol, Nopco GPI (mixture in whichfatty acid portion is Nopalcol l-O), olive oil (Conty Products Co.,contains approximately 83% oleic, 7% linoleic, 6% palmitic and 4%stearic acid), Silicone antifoam A (Dew Corning Corp, polydimethylsiloxane), Tween 80 (Atlas Powder Co., serbitan mono-oleatepolyoxyalkylene derivative) and Vegifat Y (Nopco Chem. Co., mixture offatty acids in which oleic acid predominates). Others are Lard Oil plus6% Alkaterge C, lard oil plus 1% Monsanto dcfoam AE-l, the oxazolines ofJohnson (U. S. Patent No. 2,443,825), and soya bean oil. The octadecanolor other agent used may be dissolved in mineral oil or mineral oil maybe used alone.

Typical equipment (spark plug) for automatic addition of defoamer agentto the surface of the broth is shown by Brown and Peterson (Ind. Eng.Chem. 42, 1769 (1950)). In place of the manual operation described forRuns 3 and 6 above, this type of automatic foam detection equipment maybe easily connected to the aerosolizing equipment of the presentinvention to provide injection of limited amounts of the defoameraerosol upon demand.

It is preferred that the pressure on the supply of defoamer agent beequal to or slightly less than that in the fermentation tank, which isoften kept at slightly higher than atmospheric pressure to minimizeinflux of contaminants. The gas used to vaporize the defoamer must besupplied under pressure. A pressure of -30 pounds per square inch gage(p. s. i. g.) is convenient and practical. The gas must be non-toxic tothe fermentation; air is preferred.

Many gas-atomizing nozzles (also called pneumatic atomizing nozzles) areavailable commercially and it is required only that the design and sizeof orifice produce particles of defoamer agent of diameters of less than1000 microns; it is preferred that at least 90% of the particles havediameters in the range of 1200 microns. The size of the orifice and theamount of air passed by it are obviously dependent upon the amount ofdefoamer agent required and are so selected. The quantity of agentneeded is dependent primarily upon the size of the fermentation tank.

The stream of aerosolized defoamer agent is most conveniently added tothe fermentation by joining it to the stream of sparger air after thelast filter; a separate point of entry may be used, however, providedthat suitable protection is provided against back-flooding by the mediumat times when the aerosol is not being injected into the broth. Thestream of defoamer particles in a nontoxic gas may be added to thefermenting mixture at any point substantially below the surface; it ispreferably added near the bottom of theh tank, as in the case when it ismixed with sparger air. In certain, obvious cases it is necessary thatthe gas be sterile; e. g. in the production of penicillin. The defoamerparticles may be solid or liquid or suspensions of solids in liquids.

The use of the defoamer particles of this invention enables asubstantial reduction in the consumption of defoamer agent in suchdeep-culture, aerated fermentations as those used to produce bacitracin,streptomycin, aureomycin, neomycin, terramycin, chloroamphenicol,vitamin B12, riboflavin, gluconic acid, itaconic acid and2,3-butanediol. It is of course essential that the particular defoameragent selected be effective and non-toxic for the process in which it isused.

The advantage of the use of decreased amounts of defoamer agent includenot only saving in expense but also increased ease of isolation of thefinal product by virtue of the introduction of less contaminatingmatter.

I claim:

1. In the process of producing chemicals by the deep culture, aeratedfermentation method, the step of abating and reducing foam therein whichcomprises introducing defoamer agent substantially below the surface ofthe medium as an aerosol in a stream of non-toxic gas.

2. In the process of producing antibiotics by the deep culture, aeratedfermentation method, the step of abating and reducing foam therein whichcomprises introducing defoamer agent substantially below the surface ofthe medium as an aerosol in a stream of sterile, non-toxic gas.

3. In the process of producing penicillin by the deep culture, aeratedfermentation method, the step of abating and reducing foam therein whichcomprises introducing defoamer agent substantially below the surface ofthe medium as an aerosol in a stream of sterile, non-toxic gas.

4. In the process of producing streptomycin by the deep culture, aeratedfermentation method, the step of abating and reducing foam therein whichcomprises introducing defoamer agent substantially below the surface ofthe medium as an aerosol in a stream of sterile, nontoxic gas.

5. In the process of producing chemicals by the deep culture, aeratedfermentation method, the step of abating and reducing foam therein whichcomprises introducing defoamer agent substantially below the surface ofthe medium in a stream of non-toxic gas as a dispersion of particleshaving diameters of less than 1000 microns.

6. In the process of producing antibiotics by the deep culture, aeratedfermentation method, the step of abating and reducing foam therein whichcomprises introducing defoamer agent substantially below the surface ofthe medium in a stream of sterile, non-toxic gas as a dispersion ofparticles having diameters of less than 1000 microns.

7. In a process of producing penicillin by the deep culture, aeratedfermentation method, the step of abating and reducing foam therein whichcomprises introducing defoamer agent substantially below the surface ofthe medium in a stream of sterile, non-toxic gas as a dispersion ofparticles having diameters of less than 1000 microns.

8. In the proceess of producing streptomycin by the deep culture,aerated fermentation method, the step of abating and reducing foamtherein which comprises introducing defoamer agent substantially belowthe surface of the medium in a stream of sterile, non-toxic gas as adispersion of particles having diameters of less than 1000 microns.

9. In the process of producing chemicals by the deep culture, aeratedfermentation method, the step of abating and reducing foam therein whichcomprises introducing defoamer agent substantially below the surface ofthe medium in a stream of non-toxic gas as a dispersion of particles,90% of which have diameters of less than 200 microns.

10. In the process of producing antibiotics by the deep culture, aeratedfermentation method, the step of abating and reducing foam therein whichcomprises introducing defoamer agent substantially below the surface ofthe medium in a stream of sterile, non-toxic gas as a dispersion ofparticles, 90% of which have diameters of less than 200 microns.

11. In the process of producing penicillin by the deep culture, aeratedfermentation method, the step of abating and reducing foam therein whichcomprises introducing defoamer agent substantially below the surface ofthe medium in a stream of sterile, non-toxic gas as a dispersion ofparticles, 90% of which have diameters of less than 200 microns.

12. In the process of producing streptomycin by the deep culture,aerated fermentation method, the step of abating and reducing foamtherein which comprises introducing defoamer agent substantially belowthe surface of the medium in a stream of sterile, non-toxic gas as adispersion of particles, 90% of which have diameters of less than 200microns.

13. In the process of producing chemicals by the deep culture, aeratedfermentation method, the step of abating and reducing foam therein whichcomprises introducing defoamer agent substantially below the surface ofthe medium as a spray in a non-toxic gas.

14. In the process of producing antibiotics by the deep culture, aeratedfermentation method, the step of abating and reducing foam therein whichcomprises introducing defoamer agent substantially below the surface ofthe medium as a spray in a nontoxic gas.

15. In the process of producing penicillin by the deep culture, aeratedfermentation method, the step of aba'ting and reducing foam thereinwhich comprises introducing defoamer agent substantially below thesurface of the medium as a spray in a non-toxic gas.

References Cited in the file of this patent UNITED STATES PATENTS1,792,963 Boye Feb. 17, 1931 2,408,527 Monson Oct. 1, 1946 2,443,825Johnson June 22, 1948 2,523,245 Coppock Sept. 19, 1950 2,550,450 Brownet al. Apr. 24, 1951

1. IN THE PROCESS OF PRODUCING CHEMICALS BY THE DEEP CULTURE, AERATEDFERMENTATION METHOD, THE STEP OF ABATING AND REDUCING FOAM THEREIN WHICHCOMPRISES INTRODUCING DEFOAMER AGENT SUBSTANTIALLY BELOW THE SURFACE OFTHE MEDIUM AS AN AEROSOL IN A STREAM OF NON-TOXIC GAS.